Sj"gren's syndrome (SS) is an autoimmune disease characterized by infiltration of the lacrimal glands (LGs) by lymphocytes together with their progressive destruction and finally dysfunction. The factor(s) that initiates this disease process is still not known. Since ninety percent of all patients suffering from SS are post-menopausal women, but not all postmenopausal women develop SS, we hypothesize that a decline in the serum levels of estrogen and androgen promotes an increase in the expression of LG inflammatory mediators and cell death, which becomes more severe, extensive and persistent in a predisposed genetic background. This project is relevant to the use of Hormone Replacement Therapy, and since this has become a controversial issue, it is important to evaluate the effects of sex hormones on health conditions that predominantly affect postmenopausal women. In this application we will use ovariectomized mice as a model for the post-menopausal condition. We will compare the effects of ovariectomy (OVX) between a genetically predisposed mice model of SS (NOD.B10.H2b) and control (C57BL/10) mice. The Specific Aims (SAs) of this application are: (1) To determine the expression and concentration of inflammatory mediators, lymphocytic infiltration and the type of cell death in the LGs of control and genetically predisposed mice at different time points after OVX. To carry out these experiments, we will measure the expression and concentration of the cytokines IL-12, IL-4, IL-10, IL-12 and IL-18 and the chemokines Ccl9, CXCL10 and CXCL13, previously shown to be associated with inflammatory diseases such as SS. We will perform QRT- PCR, as well as, ELISA analysis of LG homogenates at 1 day, 3 days, 7 days and 30 days after sham operation and OVX of NOD.B10.H2b and C57BL/10 mice. The type of cell death in lacrimal cells (apoptosis or necrosis) will be ascertained by light microscopy, DNA laddering, TUNEL assay and detection of active caspase 3. Histopathological analysis will be performed to evaluate the number of lymphocytic infiltration foci. Immunofluorescent analysis using antibodies against CD4 and CD8 as markers of T cells and B220 as marker of B cells will be also performed to characterize the nature of the cells involved in infiltration. (2) To test the hypothesis that androgen or estrogen treatment following OVX prevents changes in the expression and concentration of inflammatory mediators, lymphocytic infiltration and cell death described in SA1. To perform this SA, we will investigate the effects of treatment with the androgen, 5-1 dihydrotestosterone, or estrogen 172-estradiol after OVX. As long term goals we will determine the mechanism of apoptosis, the molecular mechanism of the sex hormones and determine the roles of specific cytokines and/or chemokines by generating knockout mice. These studies will allow us to define novel targets, and composite therapies for the treatment of SS and other autoimmune diseases. PUBLIC HEALTH RELEVANCE: Sj"gren's syndrome (SS) is an autoimmune disease affecting 4 million Americans that is characterized by dysfunction and destruction of lacrimal and salivary secretory tissue [1-3]. The fact that most of the patients suffering from SS are post-menopausal women clearly suggests that a decrease in circulating sex hormone (androgens and/or estrogens) levels play an important role in the initiation of SS. However, not all post-menopausal women suffer from SS. Thus, while a decrease in sex hormones alone may not be responsible for the development of autoimmune disease, abnormal hormonal levels may act synergistically with other factors, particularly genetic background, to trigger the disease and modulate its course. Thus, several factors need to be considered simultaneously to determine the correlation between these factors. Hormone replacement therapy (HRT) has become a controversial issue for post-menopausal women. A current view is that the detriments outweigh the benefits. It is important, therefore, to evaluate the effects of sex hormones on health conditions that predominantly affect post-menopausal women. These studies will give us a better understanding of the complex correlations between sex steroids, the immune system and genetics that will allow us to define novel targets and combination therapies for more efficient treatments of autoimmune diseases. [unreadable] [unreadable] [unreadable]